Angiogenesis Assessed in Tissue Microarrays of Diagnostic Follicular Lymphoma Correlates with Outcome.

Micro-vessel density (MVD) is a powerful prognostic factor in cancers but its value in haematological malignancies is more controversial. To examine its prognostic value in follicular lymphoma (FL) we assessed number and distribution of blood vessels by high-throughput tissue microarray and automated image analysis measurements in tissue microarrays (TMA) of paraffin-embedded, diagnostic lymph node biopsies taken from fifty-nine FL patients. These patients were selected from those who died from lymphoma progression less than 5 years after diagnosis (short survivor group) (n=34) and those who survived more than 15 years from diagnosis (long survivor group) (n=25). Immunohistochemistry was used in TMAs to study the number and location of vessels staining positive for the endothelial cell markers CD31 and CD34 and pericyte coverage using PDGFR. Interactive quantification using image analysis software was used to provide details of absolute numbers of vessels from each patient, as well as vessel size and their location. Image stacks of immunofluorescence stained sections were obtained using laser-scanning confocal microscopy to trace the tumour vasculature. Results demonstrated that both total vessel count and mean vessel area were significantly different between the two groups. Samples from the long survivor group were significantly more likely to have fewer (p=0.025), but larger vessels (p=0.002) than those from the short surviving group. The differences in vessel size and number were more prominent in inter-follicular vessels compared with those inside the neoplastic follicles. The smaller and more numerous blood vessels seen in the poorer prognostic sub-group likely reflects active, sprouting angiogenesis as confirmed by confocal microscopy. This study validates the use of TMAs to examine angiogenesis and demonstrates the powerful prognostic value of assessing MVD in FL. These results suggest that sprouting angiogenesis represents a therapeutic target in this disease and ongoing studies are investigating the mechanisms contributing to alteration in angiogenesis in different prognostic subgroups in FL and in transformation of this disease. The prognostic significance of MVD assessment in TMA is currently being evaluated in a validation set of FL samples.